Diversity and persistence of Helicobacter pylori

Helicobacter pylori is an exceptionally diverse bacterial species, and every infected individual harbors a unique strain, apart from the strain concordance found in some families. The infection is usually acquired during childhood and once established, persists for life unless treated with antibiotics. When infecting a new host, the bacterium causes inflammation in the gastric lining, which in most cases is asymptomatic, but may progress to chronic gastritis, gastric or duodenal ulcers or gastric cancer.

We have performed extensive genetic analyses of H. pylori to give further insight in this immense diversity with a focus on the cytotoxin associated gene pathogenicity island (cag PAI), encoding a type IV secretion system (TFSS). First we studied two subclones, one cag PAI positive and one negative, isolated from the same biopsy. These isolates differed genetically, as determined by microarray genotyping, but were more similar to each other than to any other analyzed strain. In addition, both subclones colonized germ-free transgenic Lewis B expressing mice to an equal density, but the cag PAI negative strain did not colonize conventionally raised mice, while the cag PAI positive did.

Investigating reisolates from mice infected for up to 10 months with the cag PAI positive strain revealed genetic stability of the cag PAL In the second paper we determined the nucleotide sequence of the cag PAI in four clinical isolates, two strains isolated from patients with duodenal ulcer, and two from patients with gastric cancer. These strains all harbored a functional TFSS and the overall genetic structure of this 40 kb region was similar, with some interesting exceptions. One completely new hypothetical gene, named HP0521B, was found in three of these strains. This gene was present in about half of the Swedish clinical isolates in our study. In addition, in one duodenal ulcer strain a large insertion or rearrangement in the intergenic region between HP0546 and HP0547 was found. This genomic change did not seem to affect the function of the TFSS.

The genetic composition of the H. pylori population in two individuals was examined in paper three. Subclones isolated from two time points with a nine-year interval were used. When sequencing ten loci (-6,000 bp) in three subclones from each time point, only two substitutions were found in one patient and no differences in the other. Further microarray genotyping revealed distinct differences between the subclones within the patients regardless of time of collection, indicating a potent micro-diversity. In the fourth paper we characterized a putative Nudix hydrolase, NudA, in H. pylori. The preferred substrate of NudA was AP4A, a molecule found at elevated concentrations in cells exposed to oxidative and heat stress. When challenging an isogenic mutant of nudA and a wild type with hydrogen peroxide, the mutant was less capable to survive. NudA was found to be an abundant enzyme in H. pylori, expressed at equal amounts at all stages of growth and during stress exposure.

This work has increased our understanding of the complex genetic diversity of H. pylori, and revealed the function of a Nudix hydrolase likely to be important for the persistence of this bacterium in the human stomach.

List of scientific papers

I. Bjorkholm B, Lundin A, Sillen A, Guillemin K, Salama N, Rubio C, Gordon JI, Falk P, Engstrand L (2001). Comparison of genetic divergence and fitness between two subclones of Helicobacter pylori. Infect Immun. 69(12): 7832-8.
https://pubmed.ncbi.nlm.nih.gov/11705965

II. Blomstergren A, Lundin A, Nilsson C, Engstrand L, Lundeberg J (2004). Comparative analysis of the complete cag pathogenicity island sequence in four Helicobacter pylori isolates. Gene. 328: 85-93.
https://pubmed.ncbi.nlm.nih.gov/15019987

III. Lundin A, Bjorkholm B, Kupershmidt I, Unemo M, Nilsson P, Andersson DI, Engstrand L (2004). When and where is the Helicobacter pylori genome variability generated? [Submitted]

IV. Lundin A, Nilsson C, Gerhard M, Andersson DI, Krabbe M, Engstrand L (2003). The NudA protein in the gastric pathogen Helicobacter pylori is a ubiquitous and constitutively expressed dinucleoside polyphosphate hydrolase. J Biol Chem. 278(14): 12574-8. Epub 2003 Jan 27
https://pubmed.ncbi.nlm.nih.gov/12551907